Influenza

1
Influenza

• Elysha Hussein
• Sarah Hall
• Ayesha Sattar
• Tuesday, February 25, 2003

2
Structure of Virion
100 n m
Influenza virions are SMALL. The average
eukaryotic cell diameter is 10,000 nm (10
microns), which is 100 times bigger than the
influenza virion diameter.
http//www.med.sc.edu85/pptvir2002/INFLUENZA-2002
.ppt
3
Influenza Subtypes

• Types A B
• 3 IMPs
• HA
• NA
• M2
• 8 Segments of RNA
• Responsible for epidemics pandemics

• Type C
• 1 IMP
• HEF
• Serves functions of both HA and NA
• 7 Segments of RNA
• Causes only mild infections

• Influenza strains are subtyped A, B, or C based
  on the relatedness of the matrix (M1) and
  nucleoprotein (NP) antigens
• All 3 subtypes can infect human, subtype A can
  also infect other mammals and birds
• Within each subtype, there are many variant
  strains

4
Subtype Viral Structure/Carriers
Type A
Type B

• Humans
• Swine
• Birds
• Horses
• Seals

• Humans

Type C

• Humans
• Swine

http//www-ermm.cbcu.cam.ac.uk/01002460a.pdf
5
Integral Membrane Proteins (IMP)
Hemagglutinin

• Trimeric Protein
• 500 copies per virion

Neuraminidase

• Tetrameric Protein
• 100 copies per virion

Matrix 2 (M2)

• Tetrameric Protein
• 10 copies per virion

http//www.biotech.ubc.ca/db/TEACH/BANK/PPT/flu2.p
pt
6
Fusion Schematic
Fusion Schematic
1) HA binds a cell GP at a Sialic Acid Binding
Site
http//ubik.microbiol.washington.edu/microm-pabio4
45/MM_445_lec3_2002_files/MM_445_lec3_2002.ppt
7
Fusion Schematic
Fusion Schematic
1) HA binds a cell GP at a Sialic Acid Binding
Site
Low pH
2) Clathrin-Coated pit endocytoses virion
http//ubik.microbiol.washington.edu/microm-pabio4
45/MM_445_lec3_2002_files/MM_445_lec3_2002.ppt
8
Fusion Schematic
Fusion Schematic
1) HA binds a cell GP at a Sialic Acid Binding
Site
3) Conformational Change Hydrophobic binding of
HA to vesicle membrane
Low pH
2) Clathrin-Coated pit endocytoses virion
http//ubik.microbiol.washington.edu/microm-pabio4
45/MM_445_lec3_2002_files/MM_445_lec3_2002.ppt
9
Fusion Schematic
Fusion Schematic
1) HA binds a cell GP at a Sialic Acid Binding
Site
3) Conformational Change Hydrophobic binding of
HA to vesicle membrane
Low pH
2) Clathrin-Coated pit endocytoses virion
4) RNPs are released into cytoplasm for
replication and transcription (vRNA and mRNA)
http//ubik.microbiol.washington.edu/microm-pabio4
45/MM_445_lec3_2002_files/MM_445_lec3_2002.ppt
10
Hemagglutinin (HA)

• IMP homotrimer of non-covalently linked monomers
• There are 15 variants of HA currently identified
• Precursor (HA0) is synthesized in the RER
  Golgi, then transported to the cell membrane
• Activated when cleaved into 2 chains (HA1 HA2)
  that join by disulfide bond
• HA1 is critical for initial fusion event
• Uses Sialic-acid-containing receptors on host
  cell glycoproteins. This receptor binding event
  is followed by endocytosis.

• HA2 is critical for fusion of virion w/ endosomal
  membrane
• Decrease in pH in endosome enables HA to undergo
  a confomational change that enables HA to fuse
  with the endosomal membrane

http//www.ccbb.pitt.edu/PDFFiles/150.pdf
11
HA Cleavage

• Specific cleavage site is a basic sequence of
  AAs.
• The site is conserved for specific species.
• Cleaving enzyme can determine pathogenicity of
  virus. If the enzyme is ubiquitous in cells, then
  those cells can make virulent influenza.
• Humans Argenine is present at cleavage site
• Cleaving enzyme is a tryptase called Clara
• Only produced in Clara cells, which are only
  found in upper respiratory tract
• Influenza infection is confined to this region
  of the body

12
Neuraminidase

• IMP heterotrimer
• There are 9 variants currently identified
  sequenced
• Catalyzes cleavage of aketosidic linkage between
  sialic acid and adjacent D-galactose or
  D-galactosamine
• HA binds sialic receptors, NA releases virus or
  progeny virus from receptor
• Roles in viral entry/exit
• Help virion navigate mucusal lining of respitory
  tract
• Release progeny virion from surface of host cell
• Newest Class of drugs Neuraminidase Inhibitors

13
Matrix 2

• IMP Homotetrameric
• Single pass transmembrane protein
• Roles in last 2 steps of entry process
• Facilitates membrane fusion in endosome
• Low pH in endosome activates M2 to open ion
  channel.
• Hydrogens enter virus and activate HA to undergo
  conformational change that results in membrane
  fusion with endosome
• As a consequence, RNPs are released into cytoplasm

http//www.northwestern.edu/neurobiology/faculty/p
into2/pinto_flu.pdf
14
Ribonucleoprotein Complexes (RNPs)

• After virion fuses with the endosome membrane,
  RNPs are shuttled to nucleus
• Each (-) ssRNA segment associates with 3
  polymerases and a nucleoprotein to form
  Ribonucleoprotein Complexes (RNPs)
• Replication vRNA?cRNA?vRNA
• Transcription vRNA?mRNA(?viral proteins)
• The RNA polymerase is unable to proofread
  during transcription
• This enables the virus to alter surface antigens
  and accounts for its ability to evade the immune
  system

15
Nomenclature

• 3 Subtypes, coupled with variance of the
  antigenicity of surface proteins (HA NA) and
  the long history of influenza epidemics
  necessitate a nomenclature system to catalogue
  each strain.

16
Genetic Reassortment (HA NA)
Antigenic Drift

• Minor changes in the antigenic character
• Mutation rate highest for type A, lowest for type
  C
• Most meaningful mutations occur in HA1 protein
• When 2 virions infect a cell, there are 256
  possible combinations of RNA for offspring.

http//www.biotech.ubc.ca/db/TEACH/BANK/PPT/flu2.p
pt
17
Antigenic Shift

• Phylogenic evolution that accounts for emergence
  of new strains of virus
• Immunologically distinct, novel H/N combinations
• Genetic reassortment between circulating human
  and animal strains is responsible for shifts
• Segmented genome facilitates reassortment
• Only been observed in type A, since it infects
  many species

18
Antigenic Shift 1997 Hong Kong

• H5N1 virus, harbored in chickens, infected humans
  via direct contact, only 6 casualties
• What made H5N1 strain so virulent?
• Post-mortem examination revealed high levels of
  cytokines and TNF-a.
• Indicates an innate, but not specific, immune
  response
• Hong Kong researchers suggest that this strain of
  the virus exacerbates the cytokine response,
  possibly causing toxic-shock symptoms or death

19
Antigenic Shift 1997 Hong Kong

• Webster et al Use reverse genetics to identify
  the gene responsible for increased virulence and
  immune system evasion
• Remove nonstructural (NS) gene from H5N1
• Insert this gene into benign strain
• Assess virulence of this new strain, compare to
  control
• Conclusion NS1 is critical for limiting
  antiviral effects of cytokines.
• Downregulates expression of genes involved in the
  pathway which signals the release of cytokines
• Single point mutation is responsible for making
  NS1 a better downregulator

20
Where does influenza act in the body?

• The influenza virus is a upper respiratory tract
  infection caused by one of the influenza virus
  pathogens (Type A, B, or C).
• Although it is called a respiratory disease, it
  affects the whole body, making you feel sick all
  over.

http//www.nlm.nih.gov/medlineplus/ency/imagepages
/17237.htm
21
Transmission from person-to-person by

• Tiny droplets that come from a persons mouth and
  nose when they cough and sneeze.
• Touching objects contaminated with particles from
  an infected persons nose and throat.

http//www.lungusa.org/diseases/cf02/influenza.ht
mlwhat
22
Symptoms

• Symptoms begin 1-4 days after infection.
• You can spread the flu before your symptoms start
  and 3-4 days after your symptoms appear.
• The following symptoms of the flu can vary
  depending on the type of virus, a persons age
  and overall health
• Sudden onset of chills and fever (101 103
  degrees F)
• Sore throat, dry cough
• Fatigue, malaise
• Terrible muscle aches, headaches
• Diarrhea
• Dizziness

23
Is it a cold or the flu?

• Symptoms Cold
  Flu
• Fever Rare
  Characteristic,high
• (102 104
  F),lasts 3 4 days
• Headache Rare
  Prominent
• General Aches Pains Slight Usual
  Often severe
• Fatigue Quite mild
  Can last up to 2 3 weeks
• Extreme Exhaustion Never
  Early and prominent
• Stuffy Nose Common
  Sometimes
• Sneezing Usual
  Sometimes
• Sore Throat Common
  Sometimes
• Chest Discomfort Mild to moderate
  Commoncan become
  hacking cough severe

24
Complications Superinfection

• A bacterial superinfection can develop when the
  influenza virus infects the lungs.
• The result?
• The bacteria that live in the nose and throat can
  descend to the lungs and cause bacterial
  pneumonia.
• Who is most at risk?
• People over 50, infants, those with suppressed
  immune function or chronic diseases.
• Other complications include bronchitis, sinusitis
  and ear infections.

http//www.ecureme.com/atlas/version2001/atlas.asp
25
Complications in children

• Studies show a link between the development of
  Reyes syndrome and the use of aspirin for
  relieving fevers caused by the influenza virus.
• The disease involves the CNS and the liver and
  children exhibit symptoms of drowsiness,
  persistent vomiting and change in personality.

26
Influenza outbreaks

• Outbreaks are associated with cold weather and
  therefore occur mostly in the winter months.
• A reason for this the contrast of the cold
  outdoor air and the heated indoor air can cause
  the drying of the respiratory tract tissues and
  render individuals more susceptible to
  contracting the flu.
• Outbreaks are likely to occur among individuals
  living together in settings such as nursing homes
  or among people who gather together indoors
  during the winter months.

27
Diagnosis

• Individuals with symptoms of influenza should see
  their doctor for a thorough physical exam.
• Rapid influenza tests, viral cultures, and serum
  samples can be used to confirm infection by the
  influenza virus since the symptoms of the flu are
  similar to the symptoms caused by other
  infections.

28
Rapid influenza tests

• These tests are 70 accurate for determining if
  the patient has been infected with the influenza
  virus and 90 accurate for determining the type
  of influenza pathogen.
• Examples of rapid influenza tests Directigen Flu
  A, Directigen Flu A B, Flu OIA, Quick Vue, and
  Zstat flu.
• Rapid influenza tests provide results in 24 hours
  and can be performed in the physicians office.

29
Viral Cultures

• Samples to be tested by viral cultures need to
  be collected from the first four days of
  infection.
• The viral culture can be performed from
  nasopharyngeal or throat swabs, nasal wash, or
  nasal aspirates.
• The results are made available within 3 to 10
  days.

30
Serum samples

• Blood samples can be tested for the presence of
  influenza antibody to diagnose recent infection.
  Two samples should be collected one sample
  within the first week of illness and a second
  sample 2-4 weeks later. If antibody levels
  increase from the first to the second sample,
  influenza infection likely occurred

31
How do you prevent infection?

• The only proven method for preventing influenza
  is a yearly vaccination approximately 2 weeks
  before the flu season begins.
• Since the influenza virus is subject to genetic
  mutations with the HA and NA proteins, new
  vaccines that consist of different influenza
  strains need to be developed each year.
• Every year, the vaccine is trivalent, meaning
  that it provides resistance to three strains of
  influenza viruses. The vaccine consists of 2
  influenza A virus pathogens and 1 influenza B
  pathogen.

32
Surveillance

• The global surveillance network determines which
  strains of the influenza virus will make-up the
  vaccine.
• The networks is made up of 200 WHO laboratories
  in 79 countries and 4 WHO Influenza Collaboratory
  Centers coordinate the work of the labs.
• During the course of the year, influenza viruses
  from patients are sent to these centers. The
  centers, in conjunction with the FDA Vaccines and
  Related Biological Products Advisory Committee,
  make recommendations as to the IV strains they
  expect to circulating in the next year.

33
Surveillance Contd

• After both parties agree, the vaccine is
  manufactured from inactivated viruses.

34
More on vaccination

• Each years vaccine takes about six months to
  produce, package and distribute.
• The influenza vaccine is currently produced in
  embryonated chicken eggs. Future possibilities a
  new growth medium could speed up vaccine
  production.

35
I already have the fluNow what?

• Increase liquid intake like water, juice, and
  soups.
• Get plenty of rest for the 7 to 10 days during
  which the symptoms may persist.
• Take anti-fever drugs to relieve the fever.
• Anti-viral drugs have recently been designed to
  treat the flu. If patients begin taking these
  drugs within 48 hours after their symptoms begin,
  the drugs may reduce the length of the illness by
  about 1 to 2 days.

36
Anti-viral drugs General background

• All anti-viral drugs inhibit viral replication
  but they act in different ways to achieve this.
• Drugs that are effective against influenza A
  viruses amantadine and rimantadine.
• Drugs that are effective against influenza A
  viruses and influenza B viruses zanamivir and
  oseltamivir.

http//wdhfs.state.wy.us/epiid/fluvac.htm
37
Zanamivir and Oseltamivir

• These drugs are neuraminidase inhibitors.
• They prevent the NA proteins on the surface of
  the IV from removing sialic acid from sialic
  acid-containing receptors.
• Viral budding and downstream replication of IV
  are inhibited when sialic acid remains on the
  virion membrane and host cell.
• The emerging IVs stick to the cell plasma
  membrane or other viruses since the sialic acid
  is still on the surface of the cell and the
  virion.

38
Neuraminidase inhibition
http//www.tamiflu.com/hcp/neuramin/neura_index.as
p
39
Amantadine and Rimantadine

• These drugs inhibit influenza virus A
  replication.
• They block they ion channel M2 protein which
  inhibits the delivery of IV RNPs from the
  endosomes to the cytosol.
• However, the gene that codes for M2 can mutate
  and confer resistance from these drugs.

http//www.tulane.edu/dmsander/WWW/335/Orthomyxov
iruses.html
40
Future Directions for protection

• Neirynck et al. suggest a universal vaccine for
  all influenza A viruses.
• HA and NA proteins are variant between the
  influenza A viruses, but the extracellular domain
  of the M2 protein is highly conserved.
• Neirynck et al. propose a vaccine based on the M2
  protein would protect infection by influenza A
  viruses.

41
Historically Speaking

• Influenza can be traced as far back as 400 BC
• In Hippocrates Of the Epidemics, he describes a
  cough outbreak that occurred in 412 BC in
  modern-day Turkey at the turn of the autumn
  season
• In Hippocrates Of the Epidemics, he describes a
  cough outbreak that occurred in 412 BC in
  modern-day Turkey at the turn of the autumn season

42
412 BC Outbreak

• Actual disease that affected the camp is still
  under debate but is theoretically influenza
• High communicable rate and autumn season onset
  are notable characteristics of influenza
• Death and funerals were a daily spectacle
• Miasma rising from bodies was fatal to the sick
  and the sick were fatal to the healthy
• Hostile ranks were forced to withdraw from the
  camp

43
18th Century Outbreak

• Between 1781-1782, an influenza epidemic infected
  2/3 of Romes population and ¾ of Britains
  population
• Disease spread to North America, West Indies, and
  South America
• Spread of pandemic culminated in New England, New
  York, and Nova Scotia in 1789
• 1781 marked the beginning of the 10-40 year cycle
  of influenza epidemics and pandemics

44
19th Century Outbreaks

• Asia 1829
• Spread to Indonesia by January 1831
• Russia 1830
• Spread throughout Russian and westward between
  1830 and 1831
• By November 1831, the influenza outbreak reached
  America
• Epidemics prevalent until 1851

45
19th Century Outbreaks

• After a forty year dormant cycle, Russian Flu
  pandemic occurred between 1889 and 1890
• Mostly deadly pandemic to that date (1889)
• Began in Central Asia during summer of 1889 and
  spread to Russia, China, North America, parts of
  Africa, and major Pacific Rim countries
• 500,000 750,000 mortalities worldwide
• Influenza had been regarded as a joke, but the
  medical profession finally started to realize
  its severity

46
Influenza in the spotlight

• 1900 JAMA article recognized influenza as a
  serious health threat
• Variable forms of influenza suggested
• Catarrhal type affects the respiratory or
  gastro-intestinal regions
• Neurotic type affects the cerebral, neuralgic,
  and the cardiac regions
• Blending of these types produces typhoid

47
20th Century Outbreaks

• 1918 Spanish Flu
• 1957 Asian Flu
• 1968 Hong Kong flu
• 1976 Swine Flu scare
• 1977 Russian Flu scare
• 1997 Avian Flu scare

48
1918 Spanish Flu

• Most lethal and infectious pandemic ever
• Flu first appeared in Kansas in March of 1918
• Within one week of the first reported case, the
  flu had spread to every state in the US
• Those who fell ill in the morning were dead by
  nightfall
• Those who survived symptoms of the flu often died
  of complications (such as pneumonia) caused by
  bacteria
• By April, virus spread to Europe, China, Japan,
  Africa, and South America
• Characterized as the First Wave high
  communicability, low lethality
• Despite low lethality, 800,000 worldwide had died
  by the summer

49
1918 Spanish Flu

• In late August, a second more virulent form
  emerged
• Characterized as the Main Wave
• Virus killed over 100,000 people per week in some
  US cities
• Spread throughout Europe, the Alaskan wilderness,
  and remote islands of the Pacific
• By October 1919, flu strain vanished
• At least 20,000,000 dead worldwide within 18
  months
• 850,000 Americans

50
1918 Spanish Flu

• Mortality was greater than the 4-year Black
  Death Bubonic Plague
• Mortality rate was 2.5, other epidemics had been
  0.1
• Unusually, most deaths associated with young,
  healthy adults
• Researchers isolated a wide selection of bacteria
  virus for influenza unknown
• Years later, H1NI strain found responsible for
  infection
• However, bacteria responsible for the severe
  secondary complications of pneumonia causing death

51
1957 Asian Flu

• Began in China and spread through Pacific
• H2N2 Strain responsible
• Mortality rate of 0.25
• Virus quickly identified
• Vaccine production began in May 1957
• Virus entered US and spread through school
  children
• Deaths occurred between Sept 1957-March 1958
• Highest rate of death in elderly
• 70,000 Americans dead

52
1968 Hong Kong Flu

• First detected in Hong Kong in early 1968
• H3N2 Strain responsible
• Wildly spread to US by December
• Mildest pandemic in 20th Century
• Immunity may have developed from Asian Flu
• School children were home for the holidays
• Improved medical care and antibiotics for
  secondary infections were available

53
1976 Swine Flu Scare

• Novel virus identified in Fort Dix labelled
  Killer Flu
• Thought to be related to 1918 Spanish Flu
• Mass vaccination campaign in US
• Virus never moved outside Fort Dix area
• If it had spread, it would have been much less
  deadly than the Spanish Flu

54
1977 Russian Flu Scare

• Started in northern China
• Influenza A/H1N1 responsible
• Epidemic disease in young children and young
  adults worldwide
• Persons born before 1957 had developed an
  immunity because of 1957Asian Flu
• Not considered a true pandemic because illness
  occurred primarily in children
• Virus was included in 1978-1979 vaccine

55
1997 Avian Flu Scare

• Isolated in Hong Kong
• A/H5N1 flu responsible
• Few hundred were
• infected
• 18 Hospitalized, 6 dead
• Flu did not spread from
• person to person
• Cause for concern because virus moved directly
  from chickens to people
• Pigs were NOT the intermediate host
• Chickens (1.5 million) were slaughtered
• No further spread afterwards

56
1999 Avian Flu scare

• Isolated in Hong Kong
• Influenza A/H9N2 responsible
• 2 children infected
• Pandemic was not started but incident is a cause
  for ongoing concern
• Continued presence in birds
• Ability to infect humans without intermediate
  host
• Influenza virus able to change and become more
  transmissible among people

57
Weaponization Bioterrorism

• High mutation rate
• Antigenic shifts
• Antigenic drifts
• Both changes produce new influenza virus variants
  and strains
• Strains which humans have no immunity against are
  likely to be causative agents of pandemics
• Communicability

58
If Influenza Strikes Again

• Influenzas destructive capacity resides in the
  pace and unpredictability of its virus evolution
• Can easily subvert the bodys immune response and
  outstrip societys efforts at containment
• Scenario of greatest concern for medical, public
  health, and political leaders
• Lead to a catastrophic epidemic severely taxing
  societys ability to care for the sick and dying

59
How can we prepare?

• Build capacity for care for mass casualties
• Physicians from all resources and space must be
  on hand
• Limited space sends the sick back home to further
  spread the virus
• Decentralized delivery of aid (i.e home care)
• Respect social mores relating to burial practices
• Proper treatment of the dead during an infectious
  disease emergency would require expeditious
  handling of corpses to prevent public health
  threats while avoiding dehumanizing mortuary
  practices
• Focus on developing a pneumonia vaccine, to
  prevent secondary, often fatal, infections which
  are facilitated by influenza infection.

60
How can we prepare?

• Characterize outbreak accurately and promptly
• Systematic reporting system would allow public
  health officials to keep the public informed
• For example www.cdc.gov gives a weekly influenza
  summary
• Latest reports are all available online

61
How can we prepare?

• Earn public confidence in emergency measures
• Neither support nor resistance to public health
  recommendations by the community should be taken
  for granted
• Successful plan for managing an epidemic would be
  conveying consistent and meaningful messages,
  serving audiences with diverse beliefs and
  languages, and acknowledging citizen concerns and
  grievances
• Guard against discrimination and allocate
  resources fairly
• Need to explain the disease to prevent prejudice
  that reinforces existing social schisms and
  inequalities
• Fairly allocate resources

62
References

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11. Pandemics and Pandemic Scares in the 20th Century
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